Part:BBa_K863101

Cellulose binding Domain of Cellulomonas Fimi Exoglucanse (Freiburg-Standard)

Cellulose binding domain of the [http://www.ncbi.nlm.nih.gov/nucleotide/327179207?report=genbank&log$=nucltop&blast_rank=3&RID=152ZCN0E01N Cellulomonas fimi ATCC 484 exoglucanase gene] in Standard 25 (Freiburg).

Usage and Biology

The CBDclos is an N-terminal domain quite close to the start of the protein-sequence (Figure 1). The NCBI-BLAST identified 92 amino acids (276 bases; green bar) as the protein domain. It belongs to the Cellulose Binding Module family 3 (pfam00942/cl03026; Figure 8) and is part of the very large cellulose binding protein. In which four other carbohydrate binding modules can be found, separated by hydrophobic docking interfaces for cellulase-proteins. This makes the conserved linking sequence unattractive as a Linker for fusion-proteins. For the coding sequence 12 bases (4 amino acids) more than the BLAST predicted were taken as conserved sequence upstream and 6 bases (2AS) downstream of the domain to secure natural folding of the domain.

Figure 1: protein-Blast of the Clostridium cellulovorans [http://www.ncbi.nlm.nih.gov/nuccore/M73817 cellulose binding protein gene (cbp A)]

The expressed protein would have 105 amino acids with a molecular weight of 11368.3 and a theoretic pI of 4.56. If the protein concentration is measured by optical density at 280 nm the extinction coefficients would be 15930 M-1 cm-1, assuming all pairs of Cys residues form cystines and 15930 M-1 cm-1, assuming all Cys residues are reduced. The [http://web.expasy.org/cgi-bin/protparam/protparam ExPASy Prot-Parameter-tool] predicted the estimated half-life 30 hours in mammalian reticulocytes, (in vitro) more than 20 hours in yeast (in vivo) and more than 10 hours in Escherichia coli (in vivo). It classified the protein as stable.

Number of amino acids	114
Molecular weight	11397.4	[Da]
Theoretical pI	7.85
Ext. coefficient at 280 nm	27625	[M-1 cm-1] assuming all pairs of Cys residues form cystines
Ext. coefficient at 280 nm	27500	[M-1 cm-1] assuming all Cys residues are reduced

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 4
  Illegal AgeI site found at 337
  
• 1000
  COMPATIBLE WITH RFC[1000]